1. Field of the Invention
The present invention relates to a novel purified biological response modifier having cell differentiation-induction activity which is herein referred to as Differentiation and Activation Factor and abbreviated as DAF and a process for the production thereof.
2. Description of the Prior Art
A phenomenon of cell differentiation is a process where cells capable of division and growth change into functionally matured cells responsible for the functions of an organism. For example, the processes are well known where hematopoietic stem cells constantly differentiate into matured blood cells such as leukocytes, erythrocytes, monocytes and macrophages. This differentiation and maturation process is controlled by various kinds of biological response modifiers, resulting in maintenance of the homeostasis of an organism.
Conversely it is believed that leukemia cells are derived from hematopoietic stem cells, wherein the hematopoietic stem cells are tumorized into growing cells during the normal differentiation and maturation process of the stem cells.
If such leukemia cells could be induced to redifferentiate into cells having normal functions, a new therapy of leukemia could be established on the basis of the differentiation principle. It is from such a point of view that recently, in addition to the conventional chemotherapy, radiation therapy and immunotherapy, a new type of therapy which detumorizes cancer cells into normal cells by redifferentiation of the cancer cells, has been actively investigated.
Proteinaceous substances from mice which induce myeloid leukemia cells to differentiate and detumorize into macrophage-like cells were reported as a D-factor [J. Cell. Physiol., 74, 223 (1969)]. The D-factors obtained from the culture of murine embryo cells are considered to be glycoprotein with a molecular weight of 40,000 to 50,000 [Gann, 68, 435 (1977)]. When a laboratory strain of myeloid leukemia cells from mice was implanted in a mouse and the D-factors derived from murine embryo cells were administered, the implanted myeloid leukemia cells were induced to redifferentiate into normal cells, and a life-prolongation effect on the mouse was observed [Gan to Kagaku Ryoho, Vol 8, No. 1, 8 (1981)]. Thus the literature as mentioned above suggests the possibility of cancer therapy with these differentiation-induction factors.
However, as the D-factors are proteinaceous substances derived from mice, they could cause an allergic response in a human body when administered. Therefore it is desirable to obtain a differentiation-induction factor derived from human stock. Proteinaceous differentiation-induction factors derived from human stock were produced by stimulation of the human peripheral lymphocyte with lectins, according to the J. National Cancer Institute, 67, 1225 (1981), Cancer Research, 42, 3928 (1982). However, none of the substantial properties of the differentiation-induction factors were described except that the factors were proteinaceous substances with a molecular weight of 25,000 and 40,000, as determined by gel filtration using Sephadex G-75 (Pharmacia Fine Chemicals A.B., Sweden).
Conversely differentiation-induction factors found in the supernatant of a culture of human T-cell leukemia cells were reported to be proteinaceous substances with a molecular weight of 50,000 to 60,000, as determined by acrylamide gel electrophoresis. The activity of the factors was lost by about 60% to 90% during the isolation procedures [Summary of Japan Tissue Culture Communication, 43, (1983)].
As described above, regarding the differentiation-induction factors derived from human peripheral lymphocytes and human T-cell leukemia cells, only the molecular weights have been reported and other properties of the factors have not been reported since these factors have not been obtained in an amount sufficient to allow the investigation of their properties.
As human peripheral lymphocytes can be obtained only from human blood, it is difficult to collect a large amount of the differentiation-induction factors from human peripheral lymphocytes and accordingly their industrial production is by no means easy. Also, since the maintenance of safety is expensive during the industrial culture of human T-cell leukemia cells which are infected with human T-cell leukemia virus (HTLV) and are proliferative, an economical production of the factors from human T-cell leukemia cells is very difficult.
On the other hand, macrophages are known to produce and secrete many kinds of biological response modifiers and are important for immune response. However, as human functional macrophages cannot be cultured, industrial production of these modifiers is also impossible.
Accordingly, it is desirable to establish a process for the production of a proteinaceous biological response modifier having differentiation-induction activity, wherein an established cell line capable of proliferating in vitro and not including a leukemia virus is used.